• Journal of the korean academy of Pediatric Dentistry
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• v.30 no.3
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• pp.391-405
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• 2003

Craniosynostosis, known as a premature fusion of cranial sutures, is a developmental disorder characterized by precocious differentiation and mineralization of osteoblasts in the calvarial sutures. Recent genetic studies have demonstrated that mutation in the homeobox gene Msx2 causes Boston-type human craniosynostosis. Additionally, the phenotype of Dlx5 homozygote mutant mouse presents craniofacial abnormalities including a delayed ossification of calvarial bone. Furthermore transcription of osteocalcin, a mature osteoblast marker, is reciprocally regulated by the homeodomain proteins Msx2 and Dlx5. These facts suggest important roles of osteocalcin, Msx2 and Dlx5 genes in the calvarial bone growth and suture morphogenesis. To elucidate the function of these molecules in the early morphogenesis of mouse cranial sutures, we have first analyzed by in situ hybridization the expression of osteocalcin, Msx2 and Dlx5 genes in the developing parietal bone and sagittal suture of mouse calvaria during the embryonic (E15-E18) stage. Osteocalcin mRNA was found in the periosteum of parietal bones from E15, and gradually more highly expressed with aging. Msx2 mRNA was intensely expressed in the sutural mesenchyme, osteogenic fronts and mildly expressed in the dura mater during the embryonic stage. Dlx5 mRNA was intensely expressed osteogenic fronts and the periostem of parietal bones. To further examine the upstream signaling molecules of transcription factor Msx2 and Dlx5, we have done in vitro experiments in E15.5 mouse calvarial explants. Interestingly, implantation of BMP2-, BMP4-soaked beads onto the osteogenic fronts after 48 hours organ culture induced etopic expressions of Msx2 and Dlx5 genes. On the other hand, overexpression of $TGF{\beta}1$, GDF-6, -7, FGF-2, -4 and Shh did not induce the expression of Msx2 and Dlx5. Taken together. these data indicate that transcription factor Msx2 and Dlx5 play critical roles in the calvarial bone and suture development, and that BMP siganling is involved in the osteogenesis of calvarial bones and the maintenance of cranial sutures through regulating these two transcriotpn factors. Furthermore, different expression patterns between Msx2 and Dlx5 suggest their specific functions in the osteoblast differentiation.

• Journal of Microbiology and Biotechnology
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• v.27 no.5
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• pp.1023-1031
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• 2017

The conformational change of cellular prion protein ($PrP^C$) to its misfolded counterpart, termed $PrP^{Sc}$, is mediated by a hypothesized cellular cofactor. This cofactor is believed to interact directly with certain amino acid residues of $PrP^C$. When these are mutated into cationic amino acid residues, $PrP^{Sc}$ formation and prion replication halt in a dominant negative (DN) manner, presumably due to strong binding of the cofactor to mutated $PrP^C$, designated as DN PrP mutants. Previous studies demonstrated that plasminogen and its kringle domains bind to PrP and accelerate $PrP^{Sc}$ generation. In this study, in vitro binding analysis of kringle domains of plasminogen to Q167R DN mutant PrP (PrPQ167R) was performed in parallel with the wild type (WT) and Q218K DN mutant PrP (PrPQ218K). The binding affinity of PrPQ167R was higher than that of WT PrP, but lower than that of PrPQ218K. Scatchard analysis further indicated that, like PrPQ218K and WT PrP, PrPQ167R interaction with plasminogen occurred at multiple sites, suggesting cooperativity in this interaction. Competitive binding analysis using $\small{L}$-lysine or $\small{L}$-arginine confirmed the increase of the specificity and binding affinity of the interaction as PrP acquired DN mutations. Circular dichroism spectroscopy demonstrated that the recombinant PrPs used in this study retained the ${\alpha}$-helix-rich structure. The ${\alpha}$-helix unfolding study revealed similar conformational stability for WT and DN-mutated PrPs. This study provides an additional piece of biochemical evidence concerning the interaction of plasminogen with DN mutant PrPs.

• Korean Journal of Microbiology
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• v.39 no.2
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• pp.76-82
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• 2003

To prepare evolved PHB depolymerase with increased activity for PHB or P(3HB-co-3HV) compared to the activity of the original PHB depolymerase from Alcaligenes faecalis T1, random mutation of the cloned PHB depolymerase gene was performed by using a DNA shuffling method. A library of mutated PHB depolymerase genes from A. faecalis T1 was fused to the ice nucleation protein (INP) gene from Pseudomonas syringae in pJHCl 1 and approximately 7,000 transformants were isolated. Using M9 minimal medium containing PHB or P(3HB-co-3HV) as the carbon source, mutants showing alteration in PHB depolymerase activity were selected from the transformants. The PHB depolymease activity of the transformants was confirmed by the formation of halo around colony and the turbidity decrease tests using culture supermatants. The catalytic activity of PHB depolymerase of the best mutant II-4 for PHB or P(3HB-co-13 mol% 3HV) was approximately 1.8-fold and 3.2-fold, respectively, higher than that of the original PHB depolymerase. DNA sequence analysis revealed that three amino acid residues (Ala209Val, Leu258Phe, and Asp263Thr) were substituted in II-4. From the mutational analysis, it was presumed that the substitution of amino acids near catalytic triad to more hydrophobic amino acids enhance the catalytic activity of PHB depolymerase from A. faecalis T1.

• Molecules and Cells
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• v.41 no.11
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• pp.979-992
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• 2018

Potato (Solanum tuberosum L.) is the third most important food crop, and breeding drought-tolerant varieties is vital research goal. However, detailed molecular mechanisms in response to drought stress in potatoes are not well known. In this study, we developed EMS-mutagenized potatoes that showed significant tolerance to drought stress compared to the wild-type (WT) 'Desiree' cultivar. In addition, changes to transcripts as a result of drought stress in WT and drought-tolerant (DR) plants were investigated by de novo assembly using the Illumina platform. One-week-old WT and DR plants were treated with -1.8 Mpa polyethylene glycol-8000, and total RNA was prepared from plants harvested at 0, 6, 12, 24, and 48 h for subsequent RNA sequencing. In total, 61,100 transcripts and 5,118 differentially expressed genes (DEGs) displaying up- or down-regulation were identified in pairwise comparisons of WT and DR plants following drought conditions. Transcriptome profiling showed the number of DEGs with up-regulation and down-regulation at 909, 977, 1181, 1225 and 826 between WT and DR plants at 0, 6, 12, 24, and 48 h, respectively. Results of KEGG enrichment showed that the drought tolerance mechanism of the DR plant can mainly be explained by two aspects, the 'photosynthetic-antenna protein' and 'protein processing of the endoplasmic reticulum'. We also divided eight expression patterns in four pairwise comparisons of DR plants (DR0 vs DR6, DR12, DR24, DR48) under PEG treatment. Our comprehensive transcriptome data will further enhance our understanding of the mechanisms regulating drought tolerance in tetraploid potato cultivars.

• Journal of Life Science
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• v.18 no.12
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• pp.1637-1643
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• 2008

This study has investigated whether extracellular HSP90 predisposes vascular smooth muscle cells (VSMCs) to pro-inflammatory phenotype. Exposure of rat aortic smooth muscle cells to HSP90 not only enhanced IL-6 release but also profoundly induced IL-6 transcript via promoter activation. HSP90-induced IL-6 promoter activation was suppressed by dominant-negative forms of Toll-like receptor (TLR)-4 and myeloid differentiation factor 88 (MyD88), but not by dominant-negative-forms of TLR-3 and TIR-domain-containing adapter-inducing interferon-${\beta}$ (TRIF). Curcumin, which inhibits dimerization of TLR-4, also attenuated the IL-6 induction by HSP90. Mutation at the NF-${\kappa}B$- or C/EBP-binding site in the IL-6 promoter region suppressed the promoter activation in response to HSP90. The gene delivery of $I{\kappa}B$ using recombinant adenoviruses and treatment with resveratrol, which inhibit NF-${\kappa}B$ activity, attenuated the HSP90-induced IL-6 release from VSMCs. The present study proposes that extracellular HSP90 would contribute to inflammatory reaction in the stressed vasculature by inducing IL-6 in VSMCs, and that TLR-4 and NF-${\kappa}B$ would play active roles in the process.

• Asian-Australasian Journal of Animal Sciences
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• v.30 no.7
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• pp.938-943
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• 2017

Objective: Qingyu pig, a Chinese indigenous pig breed, exhibits two types of coat colour phenotypes, including pure black and white with black spotting respectively. Melanocortin receptor 1 (MC1R) and agouti signaling protein (ASIP) are two widely reported pivotal genes that significantly affect the regulation of coat colour. The objectives of this study were to investigate whether the polymorphisms of these two genes are associated with coat colour and analyze the molecular mechanism of the coat colour separation in Qingyu pig. Methods: We studied the phenotype segregation and used polymerase chain reaction amplification and Sanger sequencing to investigate the polymorphism of MC1R and ASIP in 121 Qingyu pigs, consisting of 115 black and 6 white with black spotted pigs. Results: Coat colour of Qingyu pig is associated with the polymorphisms of MC1R but not ASIP. We only found 2 haplotypes, $E^{QY}$ and $E^{qy}$, based on the 13 observed mutations from MC1R gene. Among which, $E^{qy}$ presented a recessive inheritance mode in black spotted Qingyu pigs. Further analysis revealed a g.462-463CC insertion that caused a frameshift mutation and a premature stop codon, thus changed the first transmembrane domain completely and lost the remaining six transmembrane domains. Altogether, our results strongly support that the variety of Qingyu pig's coat colour is related to MC1R. Conclusion: Our findings indicated that black coat colour in Qingyu pig was dominant to white with black spotted phenotype and MC1R gene polymorphism was associated with coat colour separation in Qingyu pig.

• Journal of the Korean Association of Oral and Maxillofacial Surgeons
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• v.33 no.1
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• pp.46-54
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• 2007

PURPOSE Osteosarcoma occurring in the head and neck region is known as a malignant tumor that shows a relatively poor prognosis and, despite various treatments, clinicians have often been confounded by it. The existence or non-existence of the mutation of the gene $p16^{INK4a}$ has been used in prognosis assessment. In this study, author have attempted to determine whether methylation of the gene $p16^{INK4a}$ could be applied to forecast the progress of osteosarcomas in the head and neck region having been given poor prognoses in the diagnostic process and the early stage of treatment. RESEARCH SUBJECT AND METHOD Clinicopathologic investigations, immunohistochemical examinations, a methylation specific polymerase reaction (MSP) analysis, and a survival analysis were conducted on the tissues of 20 patients with mandibulofacial osteosarcoma. RESULTS Neither age, sex, size, smoking or non-smoking, nor region have showed a statistical significance with methylation or unmethylation of the gene $p16^{INK4a}$ and expression rates demonstrated by immunohisto- chemical examinations. A chi-square test indicated that recurrence inclination has no relation with the expression rate of p16 protein (p=0.6615), but it showed a statistical significance with methylation of the gene $p16^{INK4a}$ (p=0.0033). With respect to investigations of the survival rates, a Kaplan-Meier survival analysis found that the manifestation rate of p16 protein did not have an impact on survival (p=0.8864), but that the methylation of the gene $p16^{INK4a}$ resulted in significant differences in survival rates (p=0.0105). CONCLUSIONS The above results show that methylation of the gene $p16^{INK4a}$ could be one of the major factors that help determine the recurrence inclination and prognosis of osteosarcomas occurring in the head and neck region.

• Proceedings of the KOR-BRONCHOESO Conference
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• 1993.05a
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• pp.83-83
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• 1993

The nuclear phosphoprotein p53 is expressed in all normal cells and appears to function in cell cycle regulation. Abnormally high levels of the protein are found in many different types of cancer. In human cancer overexpression of p53 is associated with point mutations within highly conserved regions of p53 gene. These altered genes encode stable p53 proteins that can detected by standard immunocytochemical techniques unable to detect rapidly degraded wild-type protein. Using of a monoclonal antibody to p53 antigen, immunocytochemical analysis of 29 squamous cell carcinomas of tongue(n= 19) and tonsil(n= 10) was performed. Non-tumor nuclei showed all negative reactivity. Positive reactivity was found in 4/29(13.8%)of SCCs of tongue and tonsil. In sizes of primary tumor, the cases over 4cm showed more positive reactivity than the cases under 4cm(p < 0.05). There was no stastical correlation between the reactivity and histopathologic grades, the primary sites of tumor or the presence of cervical metastasis.

• Journal of Sericultural and Entomological Science
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• v.28 no.1
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• pp.43-53
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• 1986

Among many microbial pesticides, Bacillus thuringiensis is one of the most hopeful pesticide and some commercial products have been appearing on the market. Because these commercial products contain living spores and toxins of the organism, there is a danger that living spores of B. thuringiensis may be scattered by wind and cause a great damage in the sericulture areas. In order to avoide these risks it is desirable to select the strain which has low pathogenicity to the silkworm, and at the sometime being highly pathogenic to the pest insects. Thus this study has been carried out to acquire some basic informations about the procedure of desicable strain selection. Three strains of B. thuringiensis var. kurstaki, var. dendrolimus and var. aizawai were used for the pathogenicity test on the silkworm, Bombux mori and the fall webwarm, Hyphantria cunea. Those strains were investigated by the agarose gel electrophoresis patterns of plasmid DNA determine whether mutation had occured. Pathogenicity tests were carried out of using isolated crystal proteins and spore-crystal protein to mixtures of each strain, seperatively. In case of using spore-crystal protein mixture, the order of pathogenicity in varities of B. thuringiensis against B.mopri and H.cunea were kurstaki, aizawai, dendrolimus and kurstaki, dendrolimus, aizawai, respectively. But using isolated crystal proteins, dendrolimus had the highest toxicity to H. cunea and the lowest toxicity to B. mori among tested three strains. From the above results, dendrolimus was presumed the most desirable straing for using microbial pesticide.

• Journal of the Korea Academia-Industrial cooperation Society
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• v.20 no.8
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• pp.477-484
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• 2019

Several types of scavenger receptors, including the Collectin-Placenta 1 (CL-P1) receptorthat is present in mammals, are molecules that are expressed on the surfaces of endothelial cells, macrophages and smooth muscle cells. These molecules are cell surface glycoproteins that can be conjugated to oxidized low density lipoprotein (oxLDL). Among these molecules, the effect of quercetin on CL-P1 activation has been confirmed. Quercetin is known as an antioxidant that stops oxidation because it acts to remove free radicals that are responsible for the oxidation reaction. In this study, fragments from the transcription start site of the mouse CL-P1 gene promoter to the -500th base were cloned using DNA polymerase. These fragments were then introduced into macrophage like RAW264.7 cells and fibroblast-like NIH3T3 cells to study the effect of quercetin on the CL-P1 gene expression. As a result, we found that bases ranging from -250 to -350 in the anterior part where gene expression starts are important for producing CL-P1 protein. Among them, the DNA mutation experiments we performed confirmed that the E2F binding sites are critical for producing the CL-P1 protein? In addition, when quercetin was added to the RAW264.7 culture medium, which was a culture of adherent cells, observedthe phenomenon of the cells falling off from the surface of the culture container.